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Somatic embryogenesis and enhanced shoot organogenesis in Metabriggsia ovalifolia W. T. Wang.

Ouyang Y, Chen Y, Lü J, Teixeira da Silva JA, Zhang X, Ma G - Sci Rep (2016)

Bottom Line: Thus, somatic embryogenesis in this plant needs a high cytokinin concentration (BA; TDZ), as evidenced by histology.Somatic embryos germinated easily when left on the same media, but formed adventitious roots in two weeks on MS supplemented with 0.5 μM NAA, 0.5 μM IBA and 0.1% activated charcoal.Over 93% of plantlets survived following acclimatization and transfer to a mixture of sand and vermiculite (1:1, v/v) in trays.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, Guangzhou, 510650, China.

ABSTRACT
An efficient protocol providing a dual regeneration pathway via direct shoot organogenesis and somatic embryogenesis for an endangered species, Metabriggsia ovalifolia W. T. Wang, was established from leaf explants. When applied at 2.5 μM, the cytokinins 6-benzyladenine (BA) or thidiazuron (TDZ) and the auxins indole-3-butyric acid (IBA), α-naphthaleneacetic acid (NAA) and indole-3-acetic acid (IAA) could induce shoots when on basal Murashige and Skoog (MS) medium. BA and TDZ could induce more adventitious shoots (19.1 and 31.2/explant, respectively) than NAA (4.6/explant), IBA (5.7/explant) or IAA (6.4/explant). BA and TDZ at 5-10 μM could induce both shoots and somatic embryos. A higher concentration of TDZ (25 μM) induced only somatic embryos (39.8/explant). The same concentration of BA induced both adventitious shoots (23.6/explant) and somatic embryos (9.7/explant). Thus, somatic embryogenesis in this plant needs a high cytokinin concentration (BA; TDZ), as evidenced by histology. Somatic embryos germinated easily when left on the same media, but formed adventitious roots in two weeks on MS supplemented with 0.5 μM NAA, 0.5 μM IBA and 0.1% activated charcoal. Over 93% of plantlets survived following acclimatization and transfer to a mixture of sand and vermiculite (1:1, v/v) in trays.

No MeSH data available.


Related in: MedlinePlus

Light microscopic sections of somatic embryogenesis on induction medium supplemented with 5 μM and 25.0 μM TDZ from in vitro leaf explants of Metabriggsia ovalifolia.(a) Fresh leaf section. (b,c) Leaf explant after culture for 3 or 4 weeks, respectively showing primordial somatic embryo cell clumps (arrows) on medium supplemented with 25 μM TDZ. (d) Globular stage somatic embryos (arrows) were more visible on the leaf after culture for 5 weeks on medium supplemented with 25 μM TDZ. (e) A torpedo stage somatic embryo with a root apical meristem (black arrow) induced at a low concentration (5 μM) of TDZ. (f) Torpedo and heart stage somatic embryos (arrows) were visible on the surface of the leaf after culture for 6 weeks. (g) Heart stage somatic embryos (arrows) separated from surrounding callus after culture for 7 weeks on medium supplemented with 25 μM TDZ. Bars = 0.2 mm.
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f4: Light microscopic sections of somatic embryogenesis on induction medium supplemented with 5 μM and 25.0 μM TDZ from in vitro leaf explants of Metabriggsia ovalifolia.(a) Fresh leaf section. (b,c) Leaf explant after culture for 3 or 4 weeks, respectively showing primordial somatic embryo cell clumps (arrows) on medium supplemented with 25 μM TDZ. (d) Globular stage somatic embryos (arrows) were more visible on the leaf after culture for 5 weeks on medium supplemented with 25 μM TDZ. (e) A torpedo stage somatic embryo with a root apical meristem (black arrow) induced at a low concentration (5 μM) of TDZ. (f) Torpedo and heart stage somatic embryos (arrows) were visible on the surface of the leaf after culture for 6 weeks. (g) Heart stage somatic embryos (arrows) separated from surrounding callus after culture for 7 weeks on medium supplemented with 25 μM TDZ. Bars = 0.2 mm.

Mentions: The in vitro leaves consisted of 6–8 layers of mesophyll cells (Fig. 4a). Exposure to 25 μM TDZ for 4 weeks revealed no obvious changes to the leaf surface. In fact, from the third to fourth week, the meristematic tissue cells swelled and divided into 10–20 layers of cells. Both protuberances without somatic embryos as well as darkly stained embryogenic cell masses developed (Fig. 4b,c). Several globular somatic embryos were observed on the surface of one side of leaf explants, occasionally on both sides, after 5 weeks (Fig. 4d). Globular-shaped somatic embryos developed into heart- or torpedo-shaped somatic embryos after 6–7 weeks (Fig. 4e). Some somatic embryos separated easily from surrounding callus (Fig. 4f). On induction medium supplemented with 25 μM TDZ, no roots were induced due to the high concentration of this cytokinin. Consequently, no root apical meristems were observed. However, somatic embryos were easily separable from leaf sections (Fig. 4f), serving as another distinct difference between somatic embryos and adventitious shoots.


Somatic embryogenesis and enhanced shoot organogenesis in Metabriggsia ovalifolia W. T. Wang.

Ouyang Y, Chen Y, Lü J, Teixeira da Silva JA, Zhang X, Ma G - Sci Rep (2016)

Light microscopic sections of somatic embryogenesis on induction medium supplemented with 5 μM and 25.0 μM TDZ from in vitro leaf explants of Metabriggsia ovalifolia.(a) Fresh leaf section. (b,c) Leaf explant after culture for 3 or 4 weeks, respectively showing primordial somatic embryo cell clumps (arrows) on medium supplemented with 25 μM TDZ. (d) Globular stage somatic embryos (arrows) were more visible on the leaf after culture for 5 weeks on medium supplemented with 25 μM TDZ. (e) A torpedo stage somatic embryo with a root apical meristem (black arrow) induced at a low concentration (5 μM) of TDZ. (f) Torpedo and heart stage somatic embryos (arrows) were visible on the surface of the leaf after culture for 6 weeks. (g) Heart stage somatic embryos (arrows) separated from surrounding callus after culture for 7 weeks on medium supplemented with 25 μM TDZ. Bars = 0.2 mm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835782&req=5

f4: Light microscopic sections of somatic embryogenesis on induction medium supplemented with 5 μM and 25.0 μM TDZ from in vitro leaf explants of Metabriggsia ovalifolia.(a) Fresh leaf section. (b,c) Leaf explant after culture for 3 or 4 weeks, respectively showing primordial somatic embryo cell clumps (arrows) on medium supplemented with 25 μM TDZ. (d) Globular stage somatic embryos (arrows) were more visible on the leaf after culture for 5 weeks on medium supplemented with 25 μM TDZ. (e) A torpedo stage somatic embryo with a root apical meristem (black arrow) induced at a low concentration (5 μM) of TDZ. (f) Torpedo and heart stage somatic embryos (arrows) were visible on the surface of the leaf after culture for 6 weeks. (g) Heart stage somatic embryos (arrows) separated from surrounding callus after culture for 7 weeks on medium supplemented with 25 μM TDZ. Bars = 0.2 mm.
Mentions: The in vitro leaves consisted of 6–8 layers of mesophyll cells (Fig. 4a). Exposure to 25 μM TDZ for 4 weeks revealed no obvious changes to the leaf surface. In fact, from the third to fourth week, the meristematic tissue cells swelled and divided into 10–20 layers of cells. Both protuberances without somatic embryos as well as darkly stained embryogenic cell masses developed (Fig. 4b,c). Several globular somatic embryos were observed on the surface of one side of leaf explants, occasionally on both sides, after 5 weeks (Fig. 4d). Globular-shaped somatic embryos developed into heart- or torpedo-shaped somatic embryos after 6–7 weeks (Fig. 4e). Some somatic embryos separated easily from surrounding callus (Fig. 4f). On induction medium supplemented with 25 μM TDZ, no roots were induced due to the high concentration of this cytokinin. Consequently, no root apical meristems were observed. However, somatic embryos were easily separable from leaf sections (Fig. 4f), serving as another distinct difference between somatic embryos and adventitious shoots.

Bottom Line: Thus, somatic embryogenesis in this plant needs a high cytokinin concentration (BA; TDZ), as evidenced by histology.Somatic embryos germinated easily when left on the same media, but formed adventitious roots in two weeks on MS supplemented with 0.5 μM NAA, 0.5 μM IBA and 0.1% activated charcoal.Over 93% of plantlets survived following acclimatization and transfer to a mixture of sand and vermiculite (1:1, v/v) in trays.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, Guangzhou, 510650, China.

ABSTRACT
An efficient protocol providing a dual regeneration pathway via direct shoot organogenesis and somatic embryogenesis for an endangered species, Metabriggsia ovalifolia W. T. Wang, was established from leaf explants. When applied at 2.5 μM, the cytokinins 6-benzyladenine (BA) or thidiazuron (TDZ) and the auxins indole-3-butyric acid (IBA), α-naphthaleneacetic acid (NAA) and indole-3-acetic acid (IAA) could induce shoots when on basal Murashige and Skoog (MS) medium. BA and TDZ could induce more adventitious shoots (19.1 and 31.2/explant, respectively) than NAA (4.6/explant), IBA (5.7/explant) or IAA (6.4/explant). BA and TDZ at 5-10 μM could induce both shoots and somatic embryos. A higher concentration of TDZ (25 μM) induced only somatic embryos (39.8/explant). The same concentration of BA induced both adventitious shoots (23.6/explant) and somatic embryos (9.7/explant). Thus, somatic embryogenesis in this plant needs a high cytokinin concentration (BA; TDZ), as evidenced by histology. Somatic embryos germinated easily when left on the same media, but formed adventitious roots in two weeks on MS supplemented with 0.5 μM NAA, 0.5 μM IBA and 0.1% activated charcoal. Over 93% of plantlets survived following acclimatization and transfer to a mixture of sand and vermiculite (1:1, v/v) in trays.

No MeSH data available.


Related in: MedlinePlus